1. Field of the Invention
This invention relates to the isolation and characterization of a novel lytic peptide, cNK-2, which is derived from chicken NK lysin and which has a direct killing activity against multiple Eimeria species as well as against other apicomplexa parasites including Neospora and Cryptosporidia. 
2. Description of the Relevant Art
Avian coccidiosis is a ubiquitous intestinal protozoan infection of poultry seriously impairing the growth and feed utilization of infected animals. Conventional disease control strategies rely heavily on chemoprophylaxis resulting in high costs for the industry. However, due to the continual emergence of drug resistant strains of Eimeria, coupled with the increasing regulations and bans on the use of anticoccidial drugs in commercial poultry production, there is a need for novel approaches and alternative control strategies. Due to the complexity of the host immunity and the parasite life cycle, it has been difficult to develop new intervention strategies or recombinant vaccines against apicomplexa parasites.
NK-lysin has been characterized in mammalian as well as avian species (Andersson et al. 1995. FEBS Lett. 362:328-332; Davis et al. 2005. Vet. Immunol. Immunopathol. 105: 163-169; Endsley et al. 2004. J. Immunol. 173:2607-2614; Hong et al. 2006. Vet. Immunol. Immunopathol. 110: 339-347; Stenger et al. 1998. Science 282:121-125; Wang et al. 2006. Fish Shellfish Immunol. 20:419-426). NK-lysin in mammals, or granulysin in humans, is an antimicrobial and antitumor polypeptide expressed by NK cells and T lymphocytes. It possesses lytic activities against gram-positive and gram-negative bacteria, fungi and protozoan parasites (Andreu et al. 1999. Biochem J. 344 (Pt 3):845-849; Ernst at al. 2000.J. Immunol. 165:7102-7108; Gansert et al. 2003. J. Immunol. 170:3154-3161; Hong et al. 2008. Avian Dis. 52:302-305; Jacobs et al. 2003. Antimicrob. Agents Chemother 47:607-613; Stenger et al., supra). Antimicrobial peptides such as NK-lysin are widespread. More than 800 peptides have been identified from natural sources (Retrieved from the Internet: aps.unmc.edu/AP/main.php). However, only a very limited number of antimicrobial peptides are known to be effective against eukaryotic pathogens (Gelhaus et al. 2008. Antimicrob. Agents Chemother. 52:1713-1720).
Porcine NK-2, a shortened synthetic peptide comprising residues K39-K65 derived from the cationic core region of porcine NK-lysin with homologs in human and cattle, kills cancer cells as well as a variety of gram-negative and gram-positive bacteria, but exhibits no hemolytic or cytotoxic activity against human cells (Andra et al. 2004. Antimicrob. Agents Chemother. 48:1593-1599; Andra and Leippe. 1999. Med. Microbial. Immunol. 188:117-124; Schroder-Borm et al. 2005. FEBS Lett. 579:6128-6134). Moreover, porcine NK-2 peptide demonstrated that it disrupted the intracellular parasite Trypanosoma cruzi and intraerythrocytic Plasmodium falciparum (Gelhaus at al., supra; Jacobs et al., supra).
We have described the cloning and characterization of chicken NK-lysin, which exhibited anti-tumor cell activity against a retrovirus-transformed chicken B cell line and showed high cytotoxic activity against Eimeria acervulina and E. maxima sporozoites. However, no bactericidal activity was observed (Hong et al. 2006, 2008, supra).
There is a need for novel approaches and alternative strategies to control infections by apicomplexa parasites. Eimeria infects primarily the gut severely damaging its ability to absorb nutrition; whereas, Neospora and Cryptosporidia infect the gut of cattle and immunosuppressed humans. All three infections cause significant economic losses to the animal industry and there is no effective vaccines against them. Here, we have investigated the antimicrobial activity of synthetic chicken NK-lysin peptides against apicomplexan protozoa, in particular, E. acervulina sporozoites, Neospora caninum tachyzoites and Cryptosporidum parvum. Such peptide compositions permit the development of new treatment strategies.